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Master's Dissertation
DOI
https://doi.org/10.11606/D.42.2015.tde-27082015-191111
Document
Author
Full name
Vanessa Feitosa Viana da Silva
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2015
Supervisor
Committee
Mayer, Marcia Pinto Alves (President)
Pellizari, Vivian Helena
Sato, Maria Inês Zanoli
Title in Portuguese
Caracterização fenotípica e molecular de Escherichia coli isolodas de amostras de água do mar da região costeira de São Paulo, Brasil.
Keywords in Portuguese

Ambiente marinho
Análise de risco microbiológico
Categorias diarreiogênicas
Grupos filogenéticos
Susceptibilidade aos antibióticos
Abstract in Portuguese
A qualidade das águas marinhas destinadas a recreação de contato primário pode ser afetada por fontes de poluição pontuais e não pontuais. Escherichia coli é membro comensal da microbiota intestinal de humanos e animais endotérmicos e quando presente no ambiente marinho indica contaminação fecal recente. Embora a maioria das cepas desta espécie não seja patogênica para o homem, existem isolados virulentos e/ ou resistentes aos antibióticos. E.coli pode ser caracterizada em grupos filogenéticos - GF (A, B1, B2, C, D, E, F e clado I) e em categorias diarreiogênicas (ETEC, EPEC, EHEC, EIEC, EAEC e DAEC). Este trabalho objetivou caracterizar 99 isolados de E.coli obtidos de amostras água do mar de três regiões costeiras do estado de São Paulo com diferentes níveis de contaminação [Ubatuba (UBA), oligotrófico (n=20); Baixada Santista (BS), eutrófico (n=30) e canal de São Sebastião (CSS), mesotrófico (n=49)], frente à susceptibilidade aos antibióticos, GF, genes associados à virulência (GAV) e assim estimar os riscos microbiológicos à balneabilidade dos locais de coleta. A susceptibilidade a ampicilina (AMP), amoxicilina- ácido clavulânico (AMC), amicacina (AMI), cefotaxima (CTX), cefuroxima (CRX), ciprofloxacino (CIP), cloranfenicol (CLO), imipenema (IPM), piperacilina-tazobactam (PPT), sulfametoxazol-trimetropima (SUT) e tetraciclina (TET) foi determinada pelo método de disco difusão. A classificação em GF foi realizada por PCR utilizando duas metodologias (CLERMONT et al., 2000 e 2011). Os GAV (stx1, stx2, eae, bfpA, aggR, elt, esth, estp, invE e astA) foram pesquisados por PCR. Os resultados foram utilizados na avaliação do risco microbiológico por QMRA. Dezenove isolados foram resistentes a AMP, sendo UBA (3%), BS (33%) e CSS (16%). Dezessete isolados foram resistentes a TET: UBA (20%), BS (23%) e CSS (12%); 14 isolados foram resistentes a SUT: UBA e CSS (10%) e BS (23%). As frequências (%) dos GF (Clermont ET AL., 2000) comensais A e B1 foram, respectivamente (55 e 15 em UBA; 63 e 13 na BS; 69 e 8 no CSS); já as frequências (%) dos GF virulentos B2 e D foram, respectivamente, 5 e 25 em UBA; 7 e 16 na BS; 4 e 18 no CSS. As frequências (%) dos GF (CLERMONT et al., 2013) comensais A, B1 e C foram: respectivamente: (15, 15 e 5 em UBA; 20, 3 e 3 na BS; 2, 10 e 2 no CSS), já os grupos mais virulentos (B2, D, E e F) foram, respectivamente: (15, 15, 0 e 10 em UBA; 13, 27, 0 e 10 na BS; 10, 33, 8 e 18 no CSS); as frequências do clado I em UBA, BS e CSS foram respectivamente: (15, 23 e 12). Os GAV detectados em UBA foram: astA (15%) e bfpA (5%); BS: esth (3%) e astA (30%) e no CSS: stx1 (2%), eae (4%), estp (2%) e astA (47). Embora não tenha sido detectado risco à balneabilidade, observou-se isolados resistentes e GF virulentos em todos os pontos de coleta, dessa forma, esses ambientes devem ser conservados para assegurar a saúde ambiental e humana, além disso a QMRA não representou o risco total de todos micro-organismos e patógenos oportunistas presentes nessas regiões.
Title in English
Phenotypic and molecular characterization of Escherichia coli isolated from seawater samples from the coastal region of the state of São Paulo, Brazil.
Keywords in English
Escherichia coli
Antibiotics susceptibility
Diarrheagenic categories
Marine environment
Microbiological risk assessment
Phylogenetic groups
Abstract in English
The quality of seawater intended for bathing can be affected by sources of point and non-point pollution. In this pollutants may be Escherichia coli. E.coli is a commensal member of the intestinal tract of humans and endothermic animals, and when present in the marine environment indicates recent fecal contamination. Although most strains are harmless, there are virulent and/or resistant to antibiotics. E. coli can be evaluated by phylogenetic groups - PG (A, B1, B2, C, D, E, F, clade 1) and diarrheagenic categories (ETEC, EPEC, EHEC, EIEC, EAEC and DAEC). The aim of the present study was to characterize 99 E. coli isolated from seawater samples from three coastal regions of the state of São Paulo with different levels of contamination [Ubatuba (UBA), oligotrophic (n=20); Santos (BS), eutrophic (n=30) and São Sebastião Channel (CSS), mesotrophic (n=49)], in regard to antibiotics susceptibility, PG, virulence genes (VGs) associated with diarrheagenic pathotypes and to estimate the microbiological risks to bathing. The susceptibility to ampicillin (AMP), amoxicillin-clavulanic acid (AMC), amikacin (AMI), cefotaxime (CTX), cefuroxime (CRX), ciprofloxacin (CIP), chloramphenicol (CLO), imipenem (IPM), piperacillin-tazobactam (PPT), trimethoprim-sulfamethoxazole (SUT) and tetracycline (TET) was determined by disk diffusion. The classification in regard to GF was performed by PCR using two methodologies (2000 and 2013). The VGs (stx1, stx2, eae, bfpA, aggR, elt, esth, estp, invE and astA) were screened by PCR and the results were used to evaluate the microbiological risk by QMRA. Nineteen isolates were resistant to AMP: UBA (3%), BS (33%) and CSS (16%). Seventeen isolates were resistant to TET: UBA (20%), BS (23%) and CSS (12%); 14 isolates were resistant to the SUT: UBA and CSS (10%) and BS (23%). By the methodology of 2000, the frequencies (%) of commensals PG and B1 were, respectively, (55 and 15 in UBA; 63 and 13 in BS; 69 and 8 in CSS); while the frequencies (%) of virulent PG, B2 and D were, respectively (5 and 25 in UBA; 7 and 16 in BS; 4 and 18 in CSS). By the methodology of 2013, the frequencies (%) of PG commensals A, B1 and C were, respectively: (15, 15 and 5 in UBA; 20, 3 and 3 in BS; 2, 10 and 2 in CSS), on the other hand, the most virulent groups (B2, D, E and F) were, respectively: (15, 15, 0 and 10 in UBA; 13, 27, 0 and 10 in BS; 10, 33, 8 and 18 in CSS); clade 1 frequencies in UBA, BS and CSS were, respectively (15, 23 and 12). The VGs were detected in UBA; astA (15%) and bfpA (5%); BS: esth (3%) and astA (30%) and CSS: stx1 (2%), eae (4%), estp (2%) and astA (47). Although no microbiological hazard was detected to bathing, was observed resistant isolates and virulent PG, thereby these environments should be preserved to ensure the environmental and human health, moreover the QMRA did not represent the overall risk of all microorganisms and opportunistic pathogens present in these regions.
 
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Publishing Date
2015-08-28
 
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